DESCRIPTION: Oral HIV breastfeeding transmission (BFT) accounts for more than half of the cases of maternal-to-infant HIV transmission (MTIT). In spite of being a priority of HIV-1 research, prevention of oral HIV BFT is difficult to achieve because its mechanisms are not fully understood. Maternal virological and immunological parameters of infection, as well as innate immune factors present in milk, have been associated with oral HIV BFT. These associations are not always evident [i.e., transmission occurs from mothers with low viral loads (VLs) and vice versa, a large proportion of infants remain uninfected in spite of a long-term continuous exposure to high levels of maternal virus], pointing to the involvement of additional factors in transmission. Our preliminary studies showed that in African nonhuman primate (NHP) hosts of SIV: (i) MTIT is virtually nonexistent (<5%) (lower than the levels targeted by the WHO as a goal for achieving virtual elimination of HIV-1 MTIT); (ii) low MTIT contrasts with massive offspring SIV exposure both in utero and through BFT due to the high SIV prevalence (>80%) and high acute and chronic VLs in dams; (iii) resistance to SIV BFT is strongly associated with low levels of SIV target cells at mucosal sites of the offspring; (iv) susceptibility to experimental mucosal transmission is proportional to the availability of mucosal CCR5+ CD4+ T cells. Based on these observations, our hypothesis is that the levels of HIV/SIV target cells at the mucosal sites of the breastfed infant drive the efficacy of HIV BFT. To confirm our hypothesis, we identified two NHP models of HIV infection which differ in susceptibility to SIV BFT: (1) SIVmac-infected rhesus macaques (RMs) will serve as a model of effective oral SIV BFT and (2) SIVagm-infected African green monkeys (AGMs) as a model of resistance to oral SIV BFT. By comparing and contrasting these two animal models, we will investigate the mechanism of oral HIV/SIV BFT and explore new strategies to prevent oral HIV transmission. In addition to the use of two animal models in which exposure to breast milk has an opposite outcome, we developed: (1) a new challenge strategy consisting of dose escalation, which we demonstrated in a preliminary study to accurately model natural mucosal transmission, while efficiently infecting the animals; (2) new challenge tools, consisting of transmitted/founder infectious molecular clones (IMCs) that permit us to physiologically model SIV transmission and molecularly tagged IMCs of SIVmac and SIVagm which offer us the advantages of IMCs combined with the possibility to monitor the bottleneck of transmission; (3) a new assay for monitoring virus transmission and the diversity of the transmitted virus (real-time single genome amplification). In a step-wise approach, we will achieve the following Specific aims (SA): SA1. To test the hypothesis that the infant mucosa drives the efficacy of SIV BFT. SA1a. Dissect the relative contribution of milk vs. the levels of infant mucosal target cells in driving HIV-1 BFT. SA1b. Provide proof-of-concept data that reduction of target cell availability in the infant mucosa decreases SIV BFT rates. SA2. To identify the mucosal site of viral entry upon oral HIV/SIV BFT and to assess early mucosal changes that are associated with SIV BFT. SA2a. Perform virological dissections of the potential sites of entry. SA2b. Identify the exact site of SIV entry upon oral exposure through breastfeeding and the mucosal factors associated with oral SIV transmission. Relevance. Combined, these studies carried out in NHP species that vary in their susceptibility to SIV BFT, employing innovative interventions and new analytical tools, will delineate the factors responsible for oral SIV BFT and identify new strategies for preventing oral HIV transmission.